ABSTRACT Heart failure with preserved ejection fraction (HFpEF) is a growing health care burden especially in the aged population for which, to date, no therapy has proven to improve symptoms or survival. Neprilysin (NEP), a key enzyme that metabolizes natriuretic peptides (NPs), has recently been identified as an important prognostic biomarker and therapeutic target in HF with reduced ejection fraction but its role in HFpEF remains undefined. NEP is a cell membrane bound metalloendopeptidase and can be released from the cell surface, resulting in the formation of a circulating peptide that maintains its catalytic activity. The broad objective of this proposal is to define the importance of variation in circulating NEP in diastolic dysfunction (DD) and the development of HFpEF, and the significance of variation in NEP levels on responsiveness to the recently approved HF drug LCZ696 or EntrestoTM which functions in part as a NEP inhibitor. Our overall hypothesis is that an inverse relationship exists between circulating NEP and circulating NPs that affects the prevalence of DD/HFpEF, and variation in NEP levels alters response to EntrestoTM therapy. Our preliminary studies have demonstrated that significant variation in NEP protein expression exists due to genetic variation and that a linear relationship between NEP protein expression and enzymatic activity exists. To determine the association of circulating NEP with DD and HFpEF, we propose measuring serum NEP in Aim 1 in 1,402 residents (median age 64 years) from Olmsted County, Minnesota, who were first characterized as part of the National Institutes of Health (NIH) funded ``Prevalence of Left Ventricular Dysfunction Study'' (PAVD) (R01 HL55502) in whom the impact of aging was investigated on the development of DD and HFpEF. We will also measure serum NEP in 316 subjects with HFpEF (median age 69 years) enrolled in 2 NIH funded HFpEF clinical trials RELAX (Phosphodiesterase-5 Inhibition to Improve Clinical Status and Exercise Capacity in Diastolic Heart Failure) and NEAT-HFpEF (Nitrate's Effect on Activity Tolerance in Heart Failure With Preserved Ejection Fraction). HFpEF is a heterogeneous disease process and biomarker based clinical trials that deliver the right drug to the right patient is being increasingly recognized as a useful strategy to identify potential responders. Circulating NEP may play an important role as a companion biomarker to enhance efficacy of the key new HF drug that targets NEP, EntrestoTM. We will therefore perform a proof of concept study in Aim 2 that will assess the biomarker response (atrial, B-type, C-type NPs and cyclic guanosine monophosphate) to EntrestoTM in subjects with HFpEF who will be divided into groups with low and high NEP levels as determined in Aim 1. These studies will define the role of circulating NEP as a biomarker of not only disease presence but also progression as it relates to HFpEF in the elderly. We will also lay the foundation for the use of circulating NEP as a companion biomarker to potentially enhance the efficacy and individualize the use of the novel new drug EntrestoTM for the patient with HFpEF.